精索静脉曲张大鼠睾丸自噬对生精细胞的影响

Effect of testicular autophagy on spermatogenic cells in varicocele rats

目的:研究精索静脉曲张(VC)大鼠模型中睾丸不同水平自噬对生精细胞凋亡的影响. 方法:雄性SD大鼠54只随机分为6组:空白对照组、雷帕霉素对照组、氯喹对照组各6只,VC组、VC+雷帕霉素组、VC+氯喹组各12只.采用HE染色观察睾丸、附睾组织形态学变化,并对睾丸及附睾中精子形成情况进行评分,TUNEL染色检测生精细胞凋亡指数(AI),Western印迹检测LC3、p62、Bax、Bcl-2的表达. 结果:空白对照组、雷帕霉素对照组、氯喹对照组大鼠睾丸及附睾组织均未发生明显形态学变化,精子形成情况评分及AI亦无显著差异(P>0.05);VC组大鼠睾丸及附睾组织发生明显病理损伤,精子形成情况评分显著降低(P<0.01),AI显著升高(P<0.01),但VC+雷帕霉素组较VC组明显改善,而VC+氯喹组较VC组轻度加重.此外,与空白对照组比较,VC组自噬相关蛋白LC3(包括LC3-Ⅱ/LC3-I的比值)和促凋亡蛋白Bax表达显著增加(P<0.01),抑凋亡蛋白Bcl-2表达则显著降低(P<0.01);且VC+雷帕霉素组LC3与Bcl-2表达显著高于VC组(P<0.01),p62和Bax表达则显著低于VC组(P<0.01);而VC+氯喹组LC3与Bcl-2表达显著低于VC组(P<0.01),p62和Bax的表达则显著高于VC组(P<0.01). 结论:VC可诱导大鼠睾丸自噬和生精细胞凋亡,上调自噬可抑制生精细胞凋亡,阻滞自噬则可促进生精细胞凋亡.

Objective:To study the effect of different levels of autophagy in the testis on the apoptosis of spermatogenic cells in the rat model of varicocele(VC).Methods:We randomly divided 54 SD male rats into six groups,blank control(n=6),rapamycin control(n=6),chloroquine control(n=6),VC model control(n=12),VC+rapamycin(n=12),and VC+chloroquine(n=12).We observed the histomorphological changes of the testis and epididymis by HE staining,obtained the scores on spermatogenesis in the testis and epididymis,calculated the apoptosis index(AI)of the testicular spermatogenic cells by TUNEL,and determined the expressions of LC3-D,LC3-I,p62,Bax and Bcl-2 proteins in the testis tissue by Western blot.Results:There were no significant morphological changes in the testis and epididymis of the rats in the blank control,rapamycin control and chloroquine control groups,or significant differences in the scores on testicular and epididymal spermatogenesis and AI of the testicular.spermatogenic cells(P>0.05).The animals in the VC model control group exhibited significant pathologic al damage in the testicular and epididymal tissues,with remarkably decreased scores on spermatogenesis(P<0.01)and increased AI(P<0.01),which were markedly improved in the VC+rapamycin group and slightly aggravated in the VC+chloroquine group compared with the VC model controls.In comparison with the rats in the blank control group,those in the VC model control group showed significantly up-regulated expressions of the autophagy-related protein LC3(including the LC3-Ⅱ/LC3-Ⅰ ratio)and the pro-apoptotic protein Bax in testicular tissue(P<0.01)but down-regulated expression of the anti-apoptotic protein Bcl-2(P<0.01).The expressions of LC3 and Bcl-2 in the testis tissue were significantly higher in the VC+rapamycin(P<0.01)but lower in the VC+chloroquine group(P<0.01),while those of p62 and Bax remarkably lower in the VC+rapamycin(P<0.01)but higher in the VC+chloroquine group than in the VC model controls(P<0.01).Conclusion:Varicocele induces autophagy in the testis and apoptosis of spermatogenic cells in rats.Lp-regulating autophagy can inhibit while blocking autophagy can promote the apoptosis of spermatogenic cells.